Bioconversion of lignocellulosic biomass to bioethanol has shown environmental, economic and energetic advantages in comparison to bioethanol produced from sugar or starch. However, the pretreatment process for increasing the enzymatic accessibility and improving the digestibility of cellulose is hindered by many physical-chemical, structural and compositional factors, which make these materials difficult to be used as feed stocks for ethanol production. Lignocellulosic substrates such as, Sugarcane waste, mushroom bed waste and corn waste were pretreated by physical, chemical and biological methods. Pretreatment with NaOH (6%) released a maximum of 120% and 108% cellulose in both sugarcane waste and corn waste whereas H₂SO₄ (6%) released 84% and 84% cellulose in vegetable waste and mushroom bed waste. The pretreated substrates were analyzed by FTIR.